<2021-09-14 Tue> David biweekly meeting

projects

Project 4: Reconstitution of endocytic actin network

recent progress

  • one +Las17 sample survived all the way to imaging
    • example fluorescence region: 4.16.9EM2.1_1j_summary.png
      • learned that it's not possible to correlate against the position of the grid label
      • correlation needs to be relative to landmarks within the sample itself, which would require an irregular distribution of beads
    • EM of sections from that region:

      2021-09-14_13-44-40_sample2.1_j1_grid2a_0001.jpg
      2021-09-14_13-45-06_sample2.1_j1_grid2a other side_0009.jpg
      2021-09-14_13-45-24_sample2.1_j1_grid2a other side_0017.jpg
      2021-09-14_13-45-38_sample2.1_j1_grid2a other side_0014.jpg
      • maybe this is an actin tail? too bad we can't correlate to fluorescence
  • membranes look similar to past samples I've prepared using the old technique for EM processing
  • got trained on EM sample processing and imaging, so I have a better idea of how to design the experiment now

next steps

week of <2021-09-13 Mon>
  • prep new reactions and try to identify regions that are amenable to correlation
week of <2021-09-20 Mon>
  • determine whether correlation is possible with these samples
week of <2021-09-27 Mon>
  • if correlation is possible, determine whether there is a difference in membrane shape between high and low fluorescence beads
    • come up with some metric of scoring if so
weeks of <2021-10-04 Mon> to <2021-10-22 Fri>
  • if there are differences, collect triplicate data and do statistical test

Project 7: Theoretical membrane bending by protein LLPS

recent progress

  • finished writing up all equations for membrane-only mechanics model

    2021-09-14_11-57-34_boundaries_v1.png

    The dynamics of the \(i\)'th segment are governed by

    \begin{subequations} \begin{align} \dot r_i=-\frac{\Gamma}{\Delta A_i}\frac{dF}{dr_i}\\ \dot z_i=-\frac{\Gamma}{\Delta A_i}\frac{dF}{dz_i} \end{align} \end{subequations}

    where

    \begin{equation} F=F_H + F_\gamma +F_V\ \end{equation} \begin{equation} F_H=&2\kappa_3 \left( \left(H(r_1, z_1) - H_3 \right) \Delta A_1 + \sum_{i=2}^{3N-1} \left(H(r_i, z_i) - H_3 \right) \Delta A_i + \left(H(r_{3N}, z_{3N}) - H_3 \right) \Delta A_{3N} \right) \end{equation} \begin{equation} F_\gamma=&(\sigma-\gamma_3)\left(\Delta A_1 + \sum_{i=2}^{3N-1} \Delta A_{i} + \Delta A_{3N} - \pi r_{3N}^2\right) \end{equation} \begin{equation} F_V=&P\left(\Delta V_{1} + \sum_{i=2}^{3N-1} \Delta V_{i} + \Delta V_{3 N}\right) \end{equation}
  • starting to write up code for simulating the model

    2021-09-14_11-58-19_20210908_programming_schematic.png

next steps

short-term
  • write code to simulate model
  • add droplet to model
long-term
  • brainstorm more figure ideas
  • turn figure ideas into real figures
  • brainstorm experimental complements
    • use a GUV or SLB system and tether proteins droplets to observe membrane deformation like in yuan_membrane_2020
      • can tweak parameters like droplet volume, wetting energy, surface tension in both experiment and model
      • might even be able to change spontaneous curvature and rigidity by adding scaffold proteins
      • nanofabricated substrates might be best to initiate curvature
    • figure out a way to isolate actin clumps from sekiya-kawasaki_dynamic_2003 and stick them on synthetic membranes
    • get actin-independent CME working in live cells like they did in soheil_aghamohammadzadeh_differential_2009
      • test how multivalency affects CME dynamics
    • look into protoplast cells to reduce turgor pressure

Date: \today

Author: Max Ferrin

Email: ferrinm@berkeley.edu

Created: 2021-09-14 Tue 14:17

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